As a rule, no. Transient expression systems tend to use strong promoters such as CMV, yielding very high expression levels for a day or two, after which time, expression levels often dramatically decrease as plasmids are lost during cell division. Stable expression systems do not usually disappate over time, and may or may not achieve a high level of expression, depending on the choice of promoter and its level of function in a target cell type.

What is the time-scale of your planned experiment?

As a rule, no. Transient expression systems tend to use strong promoters such as CMV, yielding very high expression levels for a day or two, after which time, expression levels often dramatically decrease as plasmids are lost during cell division. Stable expression systems do not usually disappate over time, and may or may not achieve a high level of expression, depending on the choice of promoter and its level of function in a target cell type.

What is the time-scale of your planned experiment?

Thanks Daniel for your quick response. Based on the requirements and limitations with the experiments design, I usually run the experiments at the maximum 6 days.

I would recommend a viral vector rather than a plasmid-based approach for this time scale, at least for mammalian cells. If you are working in yeast or bacteria, these criteria may differ, so please specify your target cell type/species.

It depends on the system, the protein, the vector used, etc....

Only intrinsic characteristic of the protein which do not depend on the level of expression should remain, but then not always. Transient transfection produces very high expression but cannot always be easily controlled. One way to overcome this problem is to use inducible expression which allows high level of expression in a stable system, the level of expression can also be modulated, it brings down the cost of doing repetitive experiments and can improve reproducibility.

So from what Daniel says in his reply, even if you are using the exact same construct in the transient transfection approach as was used to construct the stable heritable line (same promoter, same enhancers, same 3' UTR, same transcription terminating sequences) you will likely see variation. For stable lines, this could be due to a number of factors like position affects due to site of integration and copy number. Similarly, transient transfection will have a range in copy number and stability (retention of the plasmid) on the cell division by cell division time timeline. It's a tricky business and needs careful controls.

As Daniel said, transient expression systems yield a high expression level for couple of days but then it disappears. Six days is quite a stretch but please tell more about your cells and your targets.

Stable expression is supposedly what it is called: you have stable expression of your gene of interest in a number of cell passages. They take quite a lot of time to develop though.

Also, I should warn you, it is not unheard when after several passages your stable cell line stop to produce your protein of interest. That is a rare event (in my experience) but very unpleasant one to discover so at initial stages of work with newly developed stable cells I like to run qPCR/Western blots from several passages to make sure that I can rely on those cells.

I doubt;

We found that stable transfection is likely to lead to cellular adaptation, a phenomenon that might be mitigated by transient transfection. See our study on Werner in JBC, 286, 10017, 2011 and some of the references therein.

Best wishes,

Albino

I dont think so , because of high expression levels

When a stable cell lines stops producing your protein of interest, it could be because of promotor methylation. You might see this with the cmv promotor used in human cells, but in most cases no problems arise. We see this effect mostly when using human cancer cells in xenograft mouse models.

In most cases in stable transfectants you will not get as high expression as in transient transfections even if you use the identical constructs. Expression in stable transfectants depends on integrations site, the number of integrations and the intracellular regulation of protein stability. Cells tend to keep levels of proteins constant. We experienced that an increase in the expression of the transfected transgene resulted in a concomitant decrease of the endogenous proteins.

You can use integrating vectors like lentivirus, they efficientely transduce primary and stable cells giving a robust expression of the interest gene.